GPR37 and GPR37L1 are orphan G protein-coupled receptors that are most abundantly expressed in the central nervous system. We assessed potential activation of GPR37 and GPR37L1 by a handful of orphan neuropeptides and found that both receptors can be activated in transfected non-neuronal cells by prosaptide, which is a 14-amino-acid peptide derived from the precursor protein prosaposin. Prosaptide is known to exert neuroprotective actions in certain areas of the brain, such as the substantia nigra, via unidentified G protein-coupled receptors, and also known to exert beneficial effects in vivo on relieving neuropathic pain and promoting remyelination in animal models of nerve damage. The overall goals of the experiments described in this exploratory R21 application are to test whether GPR37 and/or GPR37L1 mediate the ability of prosaptide to stimulate survival signaling in primary neurons and promote the survival of dopaminergic neurons in vivo. The signaling studies will focus on primary cultures of dopaminergic neurons from the ventral mesencephalon and cortical neurons, which preferentially express GPR37 vs. GPR37L1, respectively. Cultures will be prepared from wild-type mice as well as from mice that are null for expression of GPR37 (GPR37-KO) or GPR37L1 (GPR37L1-KO). The ability of prosaptide to stimulate phospho-ERK and phospho-Akt in these neurons will then be assessed in cultures prepared from the WT vs. KO mice. In addition to these experiments studying the effects of prosaptide on survival signaling, we will also challenge the neurons with insults (such as trophic factor withdrawal or treatment with the toxin MPP+) in the absence and presence of varying doses of prosaptide to assess the effects of the peptide on neuronal survival. In parallel with the experiments on cultured neurons, the whole-animal studies will assess the neuroprotective effects of prosaptide on dopaminergic neurons in the substantia nigra following treatment with the dopaminergic neurotoxin MPTP. We will employ 2 different MPTP regimens (both acute and chronic) to assess the role of GPR37 and GPR37L1 in the neuroprotective effects of prosaptide in vivo. These MPTP toxicity studies will be performed in parallel with WT, GPR37-KO and GPR37L1-KO mice. If the work described in this proposal can establish GPR37 and/or GPR37L1 as the receptor(s) that mediate the neuroprotective actions of prosaptide in primary neuronal cultures and in vivo, then future studies can focus on the discovery of small molecule agonists and/or positive allosteric modulators of these receptors that might serve as novel therapeutics in the treatment of Parkinson's Disease, stroke, neuropathic pain and myelination disorders.